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Excessive loading, as seen in chronic respiratory diseases and experimentally produced by
tracheal banding (TB) may result in injury to the myofibres of the diaphragm and hypercapnic
ventilatory failure. Neutrophils (PMNs) are primary inflammatory cells that migrate to an area of
injury in order to phagocytose cellular debris and promote the inflammatory response. During this
process, neutrophils may also amplify muscle injury. We hypothesized that neutrophil depletion
would reduce diaphragm muscle injury in an animal model where diaphragm injury was induced
by tracheal banding. Adult male Sprague-Dawley rats were randomly assigned to one of four
groups: 1. control (C), 2. tracheal banded (TB), 3. control plus neutrophil depleted (C+ND), 4.
tracheal banded plus neutrophil depleted (TB+ND). In the C+ND and TB+ND groups,
neutrophils were depleted by administering anti-rat polymorphonuclear leukocyte antisera,
beginning one day pre-surgery. After anaesthesia, the carotid artery was chronically cannulated,
esophageal pressures (Pes) were measured and a cuff was tightened around the isolated trachea,
until the Pes during tidal breathing (Pes[sub TV]) was 20% of the maximal Pes (Pes[sub Max]) during tracheal
occlusion in the TB and TB+ND groups. Arterial blood gas (ABG) samples were taken daily
from the awake animal. Three days later, following anaesthesia, in vivo measures of Pes were
repeated and then the diaphragm and trachea were excised for histological and
immunohistochemical analysis. The following parameters were investigated: (1) the presence of
neutrophils in the diaphragm measured by neutrophil specific myeloperoxidase (MPO) activity and
point counting H&E diaphragm cross-sections; (2) muscle injury as quantified by point counting
H&E stained diaphragm cross-sections; (3) the presence of EDI positive macrophages in the
diaphragm by using immunohistochemistry; and (4) ABGs measure daily post-surgery. The TB
groups were hypercapnic (PaC0₂ > 45mmHg; p<0.001) and had a decreased tracheal cross
sectional area (p<0.001). ND groups had PMN counts <0.5 million/mL. MPO activity tended to
increase indicating an increased presence of neutrophils in the diaphragm of the TB group, and C
values were similar to those in the TB+ND group. The TB group had the most abnormal and
inflamed muscle (p<0.05), whereas, the TB+ND group was similar to C values. EDI positive
macrophages per cross-sectional area was greater in the TB group compared to the C group,
indicating that the macrophage population increased in response to muscle injury. We conclude
that macrophages increase and neutrophils tend to increase in the injured diaphragm induced by
tracheal banding. Neutrophil depletion decreased diaphragm injury induced by resistive loading.

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